This invention relates to culturing of microbial, mammalian, and plant cells, and more specifically to biphasic media culture apparatus for simultaneous inoculation and culture of cells in media having differing compositions and particularly having differing physical characteristics.
In laboratory practice, it is well known in procedures for the detection and identification of infecting organisms, for example, to initially culture a specimen or sample by inoculation into a liquid nutrient medium or broth, and subsequently sub-culture from the liquid medium to a solid nutrient medium in order to isolate individual colonies of a particular organism. The isolated organisms can then be identified or further cultured.
Most commonly, the liquid medium is contained in a tube or bottle, and the solid medium, which generally has an agar base and is introduced warm or hot as a liquid and gels to a solid on cooling to the temperature at which it will be used, is separately contained in a petri dish or plate. The sub-culture from the liquid medium must be aseptically transferred to a separate dish or plate containing the solid medium. Having both the liquid and solid media in a single container allows for proliferation of infecting organisms in the liquid medium, thus simplifying their detection, and then, by washing the broth over the solid medium, subculturing for isolation can be effected without aseptic transfers being necessary.
Culture bottles or flasks containing both liquid and solid media generally comprise a sealed container including the two different nutrient materials, with the solid medium in the form of an agar slant, and so disposed that it may be positioned out of contact with the liquid medium during the incubation period. Bottles of known design, for example, have included indentations along one or more side edges parallel to one face to retain the solid medium in suitable thickness along said face, or, similarly, cleats or ribs around which the medium can solidify may be fashioned into the face to grip the solidified medium.
Culture bottles of this type are subject to several burdensome and costly manufacturing operations since they must be filled in separate aseptic filling operations. Terminal sterilization would melt the agar which could then not be repositioned and resolidified separate from the liquid. Even if the two phases of media are aseptically filled separately, the solid phase must usually be incubated in an inverted position over the liquid to keep it out of contact with the liquid. Under these conditions, even with the application of previously cited mechanical restraints, the agar may be susceptible to pulling away from the retaining structure.